The overall goal of this research program is to determine how mammalian oogenesis is regulated so that a gamete competent of fertilization and development to live young is produced. The focus of this project is on the conditions for oocyte development in vitro that promote the acquisition of a full developmental capacity. Oocytes that were grown in vitro and induced to undergo maturation were fertilized and underwent prepost-implantation development, but at a lower frequency than oocytes grown in vivo and matured in vitro. The growth and development of oocytes in vitro depend on the metabolic cooperativity between the oocytes and granulosa cells. Therefore, the first part of this proposal is based on the premise that culture conditions that promote the normal development and function of granulosa cells will increase the frequency at which oocytes grown and matured in vitro acquire a full developmental capacity. Accordingly, it is proposed to determine whether the presence of follicle-stimulating hormone (FSH), 17Beta-estradiol, and/or elements of extracellular matrix will promote the development of cultured oocyte-granulosa cell complexes isolated from preantral follicles. The effect of these substances on the development of the cultured complexes will be assessed in terms of (1) the frequency of oocyte maturation in response to gonadotropins, and (2) the frequency at which oocytes acquire full developmental capacity. Identification of substances that stimulate oocyte maturation, and definition of their mechanism of action, would provide the practical and conceptual framework for treating immature ova obtained during clinical in vitro fertilization procedures. The second part of this proposal focuses on the mechanisms by which gonadotropins induce the maturation of oocytes that have been maintained in meiotic arrest by hypoxanthine in vitro. This purine is found in murine ovarian follicular fluid at concentrations that maintain oocytes in meiotic arrest in vitro. It will be determined (A) whether gonadotropins stimulate granulosa cells to produce a substance that induces the maturation of granulosa cell-denuded oocytes in the continued presence of hypoxanthine and (b), whether gonadotropin stimulation results in a decrease in the amount of hypoxanthine contained in, or bound to, the oocytes or alters the distribution of hypoxanthine metabolites in the oocyte-granulosa cell complex.